Background: With the recent growth of interest in cell-based therapies and radiolabeled cell products, there is a need to develop more robust cell labeling and imaging methods for in vivo tracking of living cells. This study describes evaluation of a novel cell labeling approach with the positron emission tomography (PET) isotope ⁸⁹Zr (T _1/2 = 78.4 h). ⁸⁹Zr may allow PET imaging measurements for several weeks and take advantage of the high sensitivity of PET imaging. Methods: A novel cell labeling agent, ⁸⁹Zr-desferrioxamine-NCS (⁸⁹Zr-DBN), was synthesized. Mouse-derived melanoma cells (mMCs), dendritic cells (mDCs), and human mesenchymal stem cells (hMSCs) were covalently labeled with ⁸⁹Zr-DBN via the reaction between the NCS group on ⁸⁹Zr-DBN and primary amine groups present on cell surface membrane protein. The stability of the label on the cell was tested by cell efflux studies for 7 days. The effect of labeling on cellular viability was tested by proliferation, trypan blue, and cytotoxicity/apoptosis assays. The stability of label was also studied in in vivo mouse models by serial PET scans and ex vivo biodistribution following intravenous and intramyocardial injection of ⁸⁹Zr-labeled hMSCs. For comparison, imaging experiments were performed after intravenous injections of ⁸⁹Zr hydrogen phosphate (⁸⁹Zr(HPO₄)₂). Results: The labeling agent, ⁸⁹Zr-DBN, was prepared in 55% ± 5% decay-corrected radiochemical yield measured by silica gel iTLC. The cell labeling efficiency was 30% to 50% after 30 min labeling depending on cell type. Radioactivity concentrations of labeled cells of up to 0.5 MBq/10⁶ cells were achieved without a negative effect on cellular viability. Cell efflux studies showed high stability of the radiolabel out to 7 days. Myocardially delivered ⁸⁹Zr-labeled hMSCs showed retention in the myocardium, as well as redistribution to the lung, liver, and bone. Intravenously administered ⁸⁹Zr-labeled hMSCs also distributed primarily to the lung, liver, and bone, whereas intravenous ⁸⁹Zr(HPO₄)₂ distributed to the liver and bone with no activity in the lung. Thus, the in vivo stability of the radiolabel on the hMSCs was evidenced. Conclusions: We have developed a robust, general, and biostable ⁸⁹Zr-DBN-based cell labeling strategy with promise for wide applications of PET-based non-invasive in vivo cell trafficking.

18F-Tetrafluoroborate (18F-TFB) is a promising iodide analog for PET imaging of thyroid cancer and sodium/iodide symporter (NIS) reporter activity in viral therapy applications. The aim of this study was to evaluate the safety, pharmacokinetics, biodistribution, and radiation dosimetry of high-specific activity 18F-TFB in healthy human subjects. 18F-TFB was synthesized with specific activity of 3.2 ± 1.3 GBq/μmol (at the end of synthesis). Dynamic and whole-body static PET/CT scans over 4 h were performed after intravenous administration of 18F-TFB (333–407 MBq) in four female and four male healthy volunteers (35 ± 11 years old). Samples of venous blood and urine were collected over the imaging period and analyzed by ion-chromatography HPLC to determine tracer stability. Vital signs and clinical laboratory safety assays were measured to evaluate safety. 18F-TFB administration was well tolerated with no significant findings on vital signs and no clinically meaningful changes in clinical laboratory assays. Left-ventricular blood pool time-activity curves showed a multi-phasic blood clearance of 18F-radioactivity with the two rapid clearance phases over the first 20 min, followed by a slower clearance phase. HPLC analysis showed insignificant 18F-labeled metabolites in the blood and urine over the length of the study (4 h). High uptakes were seen in the thyroid, stomach, salivary glands, and bladder. Urinary clearance of 18F-TFB was prominent. Metabolic stability was evidenced by low accumulation of 18F-radioactivity in the bone. Effective doses were 0.036 mSv/MBq in males and 0.064 mSv/MBq in females (p = 0.08, not significant). This initial study in healthy human subjects showed 18F-TFB was safe and distributed in the human body similar to other iodide analogs. These data support further translational studies with 18F-TFB as NIS gene reporter and imaging biomarker for thyroid cancer and other disease processes that import iodide.

The sodium/iodide symporter (NIS) is under investigation as a reporter for noninvasive imaging of gene expression. Although ¹⁸F-tetrafluoroborate (¹⁸F-TFB, ¹⁸F-BF₄^-) has shown promise as a PET imaging probe for NIS, the current synthesis method employing isotopic exchange gives suboptimal radiochemical yield and specific activity. The aim of this study was to synthesize ¹⁸F-TFB via direct radiofluorination on boron trifluoride (BF₃) to enhance both labeling yield and specific activity, and evaluation of specific activity influence on tumor uptake. Methods: An automated synthesis of ¹⁸F-TFB was developed whereby cyclotron-produced ¹⁸F-fluoride was trapped on a quaternary methyl ammonium anion exchange (QMA) cartridge, then allowed to react with BF₃ freshly pre-formulated in petroleum ether / tetrahydrofuran (THF) (50:1). The resultant ¹⁸F-TFB product was retained on the QMA cartridge. After rinsing the cartridge with THF and water, ¹⁸F-TFB was eluted from the cartridge with isotonic saline, passing through three neutral alumina cartridges and a sterilizing filter. Preclinical imaging studies with ¹⁸F-TFB were performed in athymic mice bearing NIS-expressing C6 glioma subcutaneous xenografted tumors to determine influence of specific activity on tumor uptake. Results: Under optimized conditions, ¹⁸F-TFB was synthesized in radiochemical yield of 20.0 ± 0.7% (n = 3, uncorrected for decay) and >98% radiochemical purity in a synthesis time of 10 min. Specific activities of 8.84 ± 0.56 GBq/µmol (n = 3) were achieved from starting [¹⁸F]fluoride radioactivities of 40-44 GBq. An avid uptake of ¹⁸F-TFB was observed in human NIS (hNIS) expressing C6 glioma xenografts as well as expected NIS-mediated uptake in thyroid and stomach. There was a positive association between the uptake of ¹⁸F-TFB in hNIS-expressing tumor and specific activity. Conclusion: A rapid, practical, and high-specific activity synthesis of the NIS reporter probe ¹⁸F-TFB was achieved via direct radiofluorination on BF₃ using an automated synthesis system. The synthesis of high specific activity ¹⁸F-TFB should enable future clinical studies with hNIS gene reporter viral constructs.

Background/Objective: Noninvasive PET imaging-based assessment of PD-L1 expression is of high clinical value for better patient selection and treatment response rates to PD-L1 immunotherapies. Due to their shorter biological half-life and faster clearance from the blood pool, radiolabeled antibody fragments are an attractive alternative for imaging than their full-length IgG counterpart. This work investigated the radiosynthesis and in vitro cell uptake of anti-PD-L1-B11-nanobody radiolabeled with ⁴⁴Sc (t_1/2 = 4.04 h) as an alternative to anti-PD-L1-B11-IgG, better suited for longer half-life radioisotopes such as ⁸⁹Zr (t_1/2 = 78.41 h). Methods: The proteins were conjugated with p-SCN-Bn-DTPA and radiolabeled at room temperature with ⁴⁴Sc, achieving a radiochemical yield of a RCY of 94.8 ± 3.1% (n = 3) for [⁴⁴Sc]Sc-B11-IgG and 73.6 ± 12.1% (n = 3) for [⁴⁴Sc]Sc-B11-nanobody, before purification. Results: Significantly higher uptake in the PD-L1₊ cells than PD-L1_KO cells was observed for both probes. However, high non-specific uptake, particularly of the radiolabeled B11-nanobody, was also observed which may negatively impact its potential as a molecular imaging probe. Conclusions: Due to the high non-specific uptake in vitro, the ⁴⁴Sc radiolabeled nanobody was not progressed to further in vivo evaluation. These results should, however, not discourage future evaluations of other nanobody based probes radiolabeled with ⁴⁴Sc, due to their well-matched biological and physical half-life.